Epigenetic silencing of tumor suppressor Par-4 promotes chemoresistance in recurrent breast cancer
Nathaniel W. Mabe, … , J. Will Thompson, James V. Alvarez
Nathaniel W. Mabe, … , J. Will Thompson, James V. Alvarez
Published August 27, 2018
Citation Information: J Clin Invest. 2018;128(10):4413-4428. https://doi.org/10.1172/JCI99481.
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Research Article Oncology

Epigenetic silencing of tumor suppressor Par-4 promotes chemoresistance in recurrent breast cancer

Abstract

Tumor relapse is the leading cause of death in breast cancer, largely due to the fact that recurrent tumors are frequently resistant to chemotherapy. We previously reported that downregulation of the proapoptotic protein Par-4 promotes tumor recurrence in genetically engineered mouse models of breast cancer recurrence. In the present study, we examined the mechanism and functional significance of Par-4 downregulation in recurrent tumors. We found that epithelial-to-mesenchymal transition (EMT) promotes epigenetic silencing of Par-4 in recurrent tumors. Par-4 silencing proceeded through binding of the EMT transcription factor Twist to the Par-4 promoter, where Twist induced a unique bivalent chromatin domain. This bivalent configuration conferred plasticity at the Par-4 promoter, and Par-4 silencing could be reversed with pharmacologic inhibitors of Ezh2 and HDAC1/2. Using an epigenome editing approach to reexpress Par-4 by specifically reversing the histone modifications found in recurrent tumors, we found that Par-4 reexpression sensitized recurrent tumors to chemotherapy in vitro and in vivo. Upon reexpression, Par-4 bound to the protein phosphatase PP1, caused widespread changes in phosphorylation of cytoskeletal proteins, and cooperated with microtubule-targeting drugs to induce mitotic defects. These results identify Twist-induced epigenetic silencing of Par-4 as a targetable axis that promotes chemoresistance in recurrent breast cancer.

Authors

Nathaniel W. Mabe, Douglas B. Fox, Ryan Lupo, Amy E. Decker, Stephanie N. Phelps, J. Will Thompson, James V. Alvarez

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Figure 8

Reexpressed Par-4 alters the phosphorylation of cytoskeletal proteins and cooperates with microtubule-targeting drugs to induce mitotic defects.

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Reexpressed Par-4 alters the phosphorylation of cytoskeletal proteins an...
(A) Western blot analysis showing L106P-Par-4 expression following 16-hour treatment with either vehicle or 1 μM Shld1 in biological triplicate. (B) Phosphopeptides were enriched from lysates of recurrent tumor cells treated with vehicle or Shld1 for 16 hours and then quantified by mass spectrometry. A total of 105 phosphoproteins were significantly altered following Par-4 expression, representing 1.56% of all measured phosphoproteins. Of the 105 altered phosphoproteins, 69 were downregulated and 36 upregulated. (C) Gene ontology analysis of downregulated and upregulated phosphoproteins. Dot color represents P value and dot size indicates the number of proteins in each category relative to the total number of altered proteins. (D) Volcano plot illustrating phosphoprotein enrichment and P values for all measured phosphoproteins following 16 hours of Shld1 or vehicle. Proteins are annotated if they were significantly altered (log2 fold change >1 and P <0.05) and are associated either with actin (red) or microtubules (blue) gene ontology families. (E) Quantification of abnormal mitoses in cells from recurrent tumor cell line 2, treated with vehicle, 100 nM Shld1, 10 nM vincristine, or with Shld1 and vincristine together, as determined by live microscopy. P values were determined by 1-way ANOVA and Tukey’s post hoc analysis and are shown as compared with vehicle. Error bars denote mean ± SEM. **P < 0.01, ***P < 0.001.